Markers of oxidative stress in adipose tissue during Trypanosoma cruzi infection

The protozoan parasite Trypanosoma cruzi causes Chagas disease. Cardiac and adipose tissues are among the early targets of infection and are sites of persistent infection. In the heart and adipose tissue, T. cruzi infection results in an upregulation of pro-inflammatory mediators. In the heart, infection is associated with an increase in the markers of oxidative stress. To date, markers of oxidative stress have not been evaluated in adipose tissue in this infection. Brown and white adipose tissues were obtained from CD-1 mice infected with the Brazil strain of T. cruzi for 15, 30, and 130 days post infection. Protein carbonylation and lipid peroxidation assays were performed on these samples. There was an upregulation of these markers of oxidative stress at all time-points in both white and brown adipose tissue. Determinants of anti-oxidative stress were downregulated at similar time-points. This increase in oxidative stress during T. cruzi infection most likely has a deleterious effect on host metabolism and on the heart.     

Methanobrevibacter smithii Is the Predominant Methanogen in Patients with Constipation-Predominant IBS and Methane on Breath

Purpose

Among irritable bowel syndrome (IBS) patients, breath methane producers overwhelmingly have constipation predominance (C-IBS). Although the most common methanogen in humans is Methanobrevibacter smithii, incidence and type of methanogenic bacteria in C-IBS patients are unknown.

Methods

By use of a questionnaire and lactulose breath testing, subjects with Rome II C-IBS and methane (>3?ppm) were selected (n?=?9). The control group included subjects with IBS who had no breath methane (n?=?10). Presence of bacterial DNA was assessed in a stool sample of each subject by quantitative-PCR using universal 16S rDNA primer. M. smithii was quantified by use of a specific rpoB gene primer.

Results

M. smithii was detected in both methane and non-methane subjects. However, counts and relative proportion of M. smithii were significantly higher for methane-positive than for methane-negative subjects (1.8?×?10⁷?±?3.0?×?10⁷ vs 3.2?×?10⁵?±?7.6?×?10⁵?copies/g wet stool, P?<?0.001; and 7.1?±?6.3?% vs 0.24?±?0.47?%, P?=?0.02 respectively). The minimum threshold of M. smithii resulting in positive lactulose breath testing for methane was 4.2?×?10⁵?copies/g wet stool or 1.2?% of total stool bacteria. Finally, area-under-curve for breath methane correlated significantly with both absolute quantity and percentage of M. smithii in stool (R?=?0.76; P?<?0.001 and R?=?0.77; P?<?0.001 respectively).

Conclusions

M. smithii is the predominant methanogen in C-IBS patients with methane on breath testing. The number and proportion of M. smithii in stool correlate well with amount of breath methane.     

Phenazopyridine associated acute interstitial nephritis and review of literature

Phenazopyridine is a urinary analgesic; commonly seen side-effects of this drug include, orange discoloration of urine, methemoglobinemia, yellowish skin discoloration, hepatitis and acute renal failure. Various case reports with phenazopyridine associated acute renal failure secondary to acute tubular necrosis have been reported in the literature. Acute kidney injury in these patients is caused by either direct injury to renal tubular epithelial cells or secondary to pigment induced nephropathy from hemolytic anemia. Hypoxic injury from phenazopyridine-induced methemoglobinemia has been well documented. We report a case of biopsy proven acute interstitial nephritis, associated with therapeutic doses of phenazopyridine without any evidence of methemoglobinemia or other mechanism of renal injury. Clinicians should be aware of the toxicity of this commonly used drug and should look closely for signs of renal insufficiency. Identifying and stopping the offending medication stays as the first step, but recent studies indicate that early steroid administration improves renal recovery, as well as decreasing the risk of progression to chronic kidney disease with fibrosis and consequent permanent renal damage.     

Expression of JP-8-induced inflammatory genes in AEII cells is mediated by NF-kappaB and PARP-1

Lung epithelial cells are critical in the regulation of airway inflammation in response to environmental pollutants. Altered activation of NF-kappaB is associated with expression of several proinflammatory factors in respiratory epithelial cells in response to an insult. Here we show that a low threshold dose (8 microg/ml) of the jet fuel JP-8 induces in a rat alveolar epithelial cell line (RLE-6TN) a prolonged activation of NF-kappaB as well as the increased expression of the proinflammatory cytokines TNF-alpha and IL-8, which are regulated by NF-kappaB. The up-regulation of IL-6 mRNA in cells exposed to JP-8 appears to be a reaction of RLE-6TN cells to reduce the enhancement of proinflammatory mediators in response to the fuel. Moreover, lung tissues from rats exposed to occupational levels of JP-8 by nasal aerosol also showed dysregulated expression of TNF-alpha, IL-8, and IL-6, confirming the in vitro data. The poly(ADP-ribosyl)ation of PARP-1, a coactivator of NF-kappaB, was coincident with the prolonged activation of NF-kappaB during JP-8 treatment. These results evidenced that a persistent exposure of the airway epithelium to aromatic hydrocarbons may have deleterious effects on pulmonary function.     

Caveolin and β1-integrin coordinate angiotensinogen expression in cardiac myocytes

Background

The cardiac renin–angiotensin system (RAS) has been implicated in mediating myocyte hypertrophy and remodeling, although the biochemical mechanisms responsible for regulating the local RAS are poorly understood. Caveolin-1 (Cav-1)/Cav-3 double-knockout mice display cardiac hypertrophy, and in vitro disruption of lipid rafts/caveolae using methyl-β-cyclodextrin (MβCD) abolishes cardiac protection.

Methods

In this study, neonatal rat ventricular myocytes (NRVM) were used to determine whether lipid rafts/caveolae may be involved in the regulation of angiotensinogen (Ao) gene expression, a substrate of the RAS system.

Results

Treatment with MβCD caused a time-dependent upregulation of Ao gene expression, which was associated with differential regulation of mitogen-activated protein (MAP) kinases ERK1/2, p38 and JNK phosphorylation. JNK was highly phosphorylated shortly after MβCD treatment (2–30 min), whereas marked activation of ERK1/2 and p38 occurred much later (2–4 h). β_1D-Integrin was required for MβCD-induced activation of the MAP kinases. Pharmacologic inhibition of ERK1/2 and JNK enhanced MβCD-induced Ao gene expression, whereas p38 blockade inhibited this response. Adenovirus-mediated expression of wild-type p38α enhanced MβCD-induced Ao gene expression; conversely expression of dominant negative p38α blocked the stimulatory effects of MβCD. Expression of Cav-3 siRNA stimulated Ao gene expression, whereas overexpression of Cav-3 was inhibitory. Cav-1 and Cav-3 expression levels were found to be positively regulated by p38, but unaffected by ERK1/2 and JNK.

Conclusion

Collectively, these studies indicate that lipid rafts/caveolae couple to Ao gene expression through a mechanism that involves β₁-integrin and the differential actions of MAP kinase family members.     

Methylation of histone H3 lysine 36 enhances DNA repair by nonhomologous end-joining

Given its significant role in the maintenance of genomic stability, histone methylation has been postulated to regulate DNA repair. Histone methylation mediates localization of 53BP1 to a DNA double-strand break (DSB) during homologous recombination repair, but a role in DSB repair by nonhomologous end-joining (NHEJ) has not been defined. By screening for histone methylation after DSB induction by ionizing radiation we found that generation of dimethyl histone H3 lysine 36 (H3K36me2) was the major event. Using a novel human cell system that rapidly generates a single defined DSB in the vast majority of cells, we found that the DNA repair protein Metnase (also SETMAR), which has a SET histone methylase domain, localized to an induced DSB and directly mediated the formation of H3K36me2 near the induced DSB. This dimethylation of H3K36 improved the association of early DNA repair components, including NBS1 and Ku70, with the induced DSB, and enhanced DSB repair. In addition, expression of JHDM1a (an H3K36me2 demethylase) or histone H3 in which K36 was mutated to A36 or R36 to prevent H3K36me2 formation decreased the association of early NHEJ repair components with an induced DSB and decreased DSB repair. Thus, these experiments define a histone methylation event that enhances DNA DSB repair by NHEJ.     

SCAN! A pharmacy-based,sun safety feasibility study

ObjectivesSkin cancer is the most common form of cancer, and individuals from the medically underserved Appalachian region are at elevated risks for cancer morbidity and mortality. Skin cancer can be prevented by decreasing ultraviolet light exposure (sunscreen sun protection factor 30, shade, clothing, sunglasses, hats) and can be caught at an early treatable stage through a routine skin examination. The Skin Cancer Awareness Now! (SCAN!) pilot project promoted skin cancer prevention and screening in community pharmacies, using a dynamic communication model. The objectives of the study were to understand (1) the feasibility of the SCAN! and (2) the preliminary impact of the SCAN!MethodsWe conducted pre- and postintervention surveys of the SCAN!, a student pharmacist–led or pharmacy resident–led intervention in community pharmacies (n = 3).ResultsParticipants (n = 90) had a mean age of 43.8 (SD= 18.4) years, were predominantly white (92.1%), without a college degree (65.6%), and had an average family income in the range of $25,000-$49,999, with approximately 16% falling below the poverty level. To begin, the SCAN! scored highly in attention (mean = 5.8), liking (mean = 6.1), comprehension (mean = 6.7), and intentions to be sun safe (mean = 6.0). Most improved in their knowledge of the amount of sunscreen needed per application for sun safety (66%, P < 0.01) and of melanoma features from pre- and postintervention (39%, P < 0.01). A multivariate analysis of variance indicated that knowledge and intentions improved (all P’s < 0.01). Interaction effects indicated that improvements in knowledge were greater for those in the rural pharmacy (P = 0.03), and improvements in perceived importance were greater for those in urban pharmacies (P = 0.01).ConclusionThe SCAN! intervention was well received by the population. Our study provides evidence that community pharmacy is a novel venue for skin cancer prevention interventions, particularly for rural, medically underserved populations.     

Alterations to the Gut Microbiome Impair Bone Strength and Tissue Material Properties

Alterations in the gut microbiome have been associated with changes in bone mass and microstructure, but the effects of the microbiome on bone biomechanical properties are not known. Here we examined bone strength under two conditions of altered microbiota: (1) an inbred mouse strain known to develop an altered gut microbiome due to deficits in the immune system (the Toll‐like receptor 5–deficient mouse [TLR5KO]); and (2) disruption of the gut microbiota (ΔMicrobiota) through chronic treatment with selected antibiotics (ampicillin and neomycin). The bone phenotypes of TLR5KO and WT (C57Bl/6) mice were examined after disruption of the microbiota from 4 weeks to 16 weeks of age as well as without treatment (n = 7 to 16/group, 39 animals total). Femur bending strength was less in ΔMicrobiota mice than in untreated animals and the reduction in strength was not fully explained by differences in bone cross‐sectional geometry, implicating impaired bone tissue material properties. Small differences in whole‐bone bending strength were observed between WT and TLR5KO mice after accounting for differences in bone morphology. No differences in trabecular bone volume fraction were associated with genotype or disruption of gut microbiota. Treatment altered the gut microbiota by depleting organisms from the phyla Bacteroidetes and enriching for Proteobacteria, as determined from sequencing of fecal 16S rRNA genes. Differences in splenic immune cell populations were also observed; B and T cell populations were depleted in TLR5KO mice and in ΔMicrobiota mice (p < 0.001), suggesting an association between alterations in bone tissue material properties and immune cell populations. We conclude that alterations in the gut microbiota for extended periods during growth may lead to impaired whole‐bone mechanical properties in ways that are not explained by bone geometry. © 2017 American Society for Bone and Mineral Research.